Ochrobactrum anthropi

D-alanine (peptides) DL-alanine (peptide) amide(s) D-amino acid arninopeptidase Ochrobactrum anthropi, Bhodococcus erythropolis 174... 

Racemic a-amino amides and a-hydroxy amides have been hydrolyzed enantio-selectively by amidases. Both L-selective and o-selective amidases are known. For example, a purified L-selective amidase from Ochrobactrum anthropi combines a very broad substrate specificity with a high enantioselectivity on a-hydrogen and a,a-disubstituted a-amino acid amides, a-hydroxyacid amides, and a-N-hydroxya-mino acid amides [102]. A racemase (a-amino-e-caprolactam racemase, EC 5.1.1.15) converts the o-aminopeptidase-catalyzed hydrolysis of a-amino acid amides into a DKR (Figure 6.38) [103]. 

An amidase from Ochrobactrum anthropi strain NCIMB 40321 has a wide snbstrate versatility for L-amides, primarily those with an a-amino gronp (Sonke et al. 2005), while the condensation prodnct of nrea and formaldehyde H2N-[CONH-CH2NH] -CONH2 is hydrolyzed by another strain of 0. anthropi (Jahns et al. 1997). 

Jahns T, R Schepp, H Kalytwasser (1997) Purification and characterization of an enzyme from a strain of Ochrobactrum anthropi that degrades condensation products of urea and formaldehyde (ureaform). Can J Microbiol AE. 1111-1117. 

Sonke T, S Ernste, RF Tandler, B Kaptein, WPH Peeters, FBI van Assema, MG Wubbolts, HE Schoemaker (2005) L-selective amidase with extremely broad substrate specificity from Ochrobactrum anthropi NCIMB 40321. Appl Environ Microbiol 71 7971-7963. 

Bacillus thuringiensis subsp. kurstaki (Btk), Agrobacterium sp. strain CP4, Ochrobactrum anthropi Bacillus thuringiensis subsp. 

Boesten, W.H.J., Raemakers-Franken, PC., Sonke, T. et al. (2003) Protein and cDNA sequences of -//- -amino acid amide racemases cloned from Ochrobactrum anthropi and Arthrobacter nicotianae, W02003106691. 

L-Alanine amide (S)-8 was converted to D-alanine (R)-9 in excellent yield and enantiomeric excess by incubation of the substrate with a-amino-e-caprolactam racemase from Achromohacter obae and D-aminopeptidase from Ochrobactrum anthropi (Scheme 2.5) [7]. 

A. L. L. Duchateau, and J. Kamphuis, Ochrobactrum anthropi NCIMB 40321 a new biocatalyst with broad-spectrum L-specific amidase activity, Appl. Microbiol. Biotechnol. 1993, 39, 296-300. 

Recently it was reported that an a-amino-e-caprolactam racemase from Achro-mobacter obae can racemise a-amino acid amides efficiently. In combination with a D-amino acid amidase from Ochrobactrum anthropi L-alanine amide could be converted into D-alanine. This tour de force demonstrates the power of the racemase [84]. If racemic amide is used as a starting material the application of this racemase in combination with a d- or L-amidase allows the preparation of 100% d- or L-amino acid, a dynamic kinetic resolution instead of DSM s kinetic resolution (Scheme 6.24). 

Table 12.2-2. Substrate specificty of Ochrobactrum anthropi cells. L-selective hydrolysis3 of amides by Ochrobactrum anthropi.

Synthesis of (S)- and (/ )-a-amino acids using nitrile hydratase (NHase) from Rhodococcus opacus, a-amino-e-caprolactam racemase (ACL) from Achromobacter obae, and different amidases (amidase 1, o-amino-peptidase from Ochrobactrum anthropi amidase 2, i-amino acid amidase from Brevundimonas diminuta) [111, 112]. / =CH3, CH CH, CH3CH(CH3), CH3CH(CH3)CH2. 

Since o-stereospecific amino acid amides and peptide hydrolases were previously unknown and were not targets of enz3unology, we started to screen for these enzymes and subsequently discovered three kinds that exhibited D-stereoselectivities for o-amino acid derivatives o-aminopeptidase [4], o-amino acid amidase [5], and alkaline o-peptidase [6] (Table 19.1). Ochrobactrum anthropi Cl-38 was isolated through an enridiment culture technique as a utilizer of o-alanine amide (o-Ala-NH ) as the sole... 

D-Aminopeptidase (DAP) Ochrobactrum anthropi Cl-38 Ser Dynamic kinetic resolution, peptide bond formation... 

Ochrobactrum anthropi SV3 was isolated as a D-valine amide degrader after a 4-month acclimation of bacterial culture. The strain produced an enzyme that was characterized as o-amino acid amidase (DaaA) [5], Furthermore, Bacillus cereus DF4-B excreted alkaline o-peptidase (ADP), which acted on the synthehc oligopeptide D-phenylalanine tetramer (o-Phe). The enzyme was characterized as the first endopeptidase acting on D-amino acid-containing peptides by recognizing the second amino acid from its N-terminus under alkaline conditions [6]. 

Komeda, H. and Asano, Y, "Gene cloning, nucleotide sequencing, and purification and characterization of the D-stereospecific amino-acid amidase from Ochrobactrum anthropi SV3." Eur.. Biochem., 267, 2028-2036 (2000). 

Gilles, C. B., Remaut, H., Villeret, V., Prang4 T, Fanuel, L., Delmarcelle, M., Joris, B., Frere, J. M., and Beeuman, J. V., "Crystal structure of a u-aminopeptidase from Ochrobactrum anthropi, a new member of the penicillin-recognizing enzyme family." Structure, 8, 971-980 (2000). 

Okazaki, S., Suzuki, A., Komeda, H., Yamaguchi, S., Asano, Y, and Yamane, T, "Crystal structure and functional characterization of a u-stereospecific amino acid amidase from Ochrobactrum anthropi SV3, a new member of the penicillin-recognizing proteins." /. Mol. Biol, 368, 79-91 (2007). 

Asano, Y., Mori, T., Hanamoto, S., Kato, Y, and Nakazawa, A., "A new D-stereospecific amino acid amidase from Ochrobactrum anthropi." Biochem. Biophys. Res. Commun., 162, 470-474 (1989). 

Recently an -transaminase from Ochrobactrum anthropi has been isolated and surprisingly showed no product inhibition by acetophenone [75]. Various ketones shown in Table 29.3 were examined. No reaction limitation for the transaminase from O. anthropi and a residual enzyme activity up to 100% were detected, compared to the transaminase from P. denitrificans showing no residual activity at the same product concentrations. 

TABLE 29.3 Ketone Inhibitor tested with the (o-Transaminase from Ochrobactrum anthropi ... 

Y. Zuo, D. Xing, J.M. Regan and B.E. Logan, Isolation of the exoelectrogenic bacterium Ochrobactrum anthropi YZ-1 by using a U-tube microbial fuel cell, Appl. Environ. Microbiol. 74, 2008,3130-3137. 

In this chapter we describe the DSM aminoamidase processes in more detail. Three different enzymatic resolution routes have been developed for the preparation of natural and synthetic amino acids using biocatalysts from different origin, i.e.. Pseudomonas pu-tida, Mycobacterium neoaurum, and Ochrobactrum anthropi. Scope and limitations and enzyme characterization of these amidases will be presented together with some specific examples. In addition, the use of some of these amino acids in peptide sjmthesis, catalytic asymmetric synthesis, and further synthetic transformations will be given. 

By this procedure three microorganisms could be identified that hydrolyzed DL-mandelic acid amide completely L-enantiospecific. The most interesting one of these three strains, based on its high cellular activity, could be identified as an Ochrobactrum anthropi strain, and this strain has been deposited at the NCIMB culture collection as NCIMB 40321. 

L-a-Amino acids have been prepared by the resolution of racemic a-amino acid amide by the L-specific aminopeptidase from Pseudomonas putida ATCC 12633 [7]. Enzyme from R putida ATCC 12633 cannot be used to resolve a-alkyl-substituted amino acid amides 103. Aminoamidase from Mycobacterium neoaurum ATCC 25795 has been used in the preparation of L-a-alkyl amino acid 104 (Fig. 34) and D-amide of a-alkyl-substituted amino acids by enzjmaatic resolution process using racemic a-alkyl amino acid amide as a substrate [169,179]. Amidase from Ochrobactrum anthropi catalyzed the resolution of a,a-disubsituted amino acids, iV-hydroxy amino acids, and a-hydroxy acid amides. The resolution process could lead to the production of chiral amino acids or amides in 50% yield. Recently, amino acid racemases have been used to get 100% yield of chiral amino acids [179]. 

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